*2.1. Parasite and Culture Condition*

*L. donovani* field isolate (K133WT), earlier derived from bone marrow aspirates of a VL patient and cryopreserved in a lab, was revived and propagated in medium M199 (Sigma-Aldrich, St. Louis, MO, USA) supplemented with 10% heat-inactivated fetal bovine serum (HI FBS, Gibco, Waltham, MA, USA), 100 IU/mL penicillin G, and 100 mg/mL streptomycin at 26 ◦C. The isolate was exposed to increasing concentrations (up to 50 µM) of artesunate drug (Sigma Aldrich, St. Louis, MO, USA) to obtain experimental artesunate-resistant parasites, which were designated as K133AS-R. The susceptibility of K133WT and K133AS-R parasites towards artesunate was determined, which showed that there was a 3.73-fold increase in the mean IC<sup>50</sup> (50% inhibitory concentration) of K133AS-R parasites at the promastigote stage, with a value of 78.63 ± 9.17 µM vs. 21.08 ± 3.15 µM, and a >3-fold increase in the mean IC<sup>50</sup> at the amastigote stage, with a value 73.09 ± 1.14 µM vs. 21.62 ± 3.24 µM for K133AS-R vs. K133WT isolates. This was reported in our previous study [41].
